Neuronal death is a major devastating consequence of status epilepticus. Seizure-induced neuronal death itself likely underlies status epilepticus-induced neurological deficits such as memory loss and may also contribute to a worsening of the epileptic condition. The mechanisms of seizure-induced neuronal death remain obscure. Understanding these mechanisms in molecular terms may provide novel therapeutic approaches aimed at preventing seizure-induced neuronal death. It is hypothesized that prolonged seizure activity kills vulnerable neurons by an oxidative mechanism involving the formation of superoxide (O2-) radicals. The hypothesis predicts that overproduction of O2- and related reactive species oxidizes critical cellular targets (i.e. proteins, lipids and DNA), ultimately resulting in neuronal death. Seizure-induced O2- production is hypothesized to arise from the mitochondria via the electron transport chain and the extracellular space via activation of the NADPH oxidase complex. Using the kainic acid model of epilepsy and a diversity of biochemical, molecular biological and analytical tools and techniques, the goal of this proposal is to define the role of O2- radicals in seizure induced neuronal death. Specific Aim 1 will determine if seizures produce oxidative damage and if this correlates with subsequent neuronal death. Aconitase inactivation (an index of O2- production), F2-isoprostanes (a non-enzymatic product of lipid peroxidation) and oxidized bases of DNA will be used as markers of oxidant damage to proteins, lipids and DNA respectively. Metalloporphyrin superoxide dismutase (SOD) mimetics that eliminate O2- and related reactive species will be tested for their ability to protect against oxidative damage and neuronal death. Specific Aim 2 will determine if compartmentalized O2- production modulates seizure-induced neuronal death. Genetically modified mice that over express or lack endogenous SOD1, SOD2 and SOD3 will be used to define the roles of cytosolic, mitochondrial and extracellular O2-, respectively. Specific Aim 3 will determine the roles of the mitochondrial electron transport chain and the plasma membrane NADPH oxidase as two potential cellular sources of seizure-induced O2- production. Polarographic studies in intact and submitochondrial particles as well as oxidative phosphorylation enzymology will be employed to determine the site(s) of mitochondrial O2- generation. These studies can advance our understanding of seizure-induced neuronal death and suggest novel therapeutic strategies for rescuing neurons in the context of status epilepticus in humans